Several neuropeptides are involved in mediating inflammatory and immuno responses peripherally and in conveying nociceptive information centrally. Substance P (SP), calcitonin gene-related peptide (CGRP) and galanin (GAL), for example, are synthesized and released by a subpopulation of primary afferent neurons thought to be important in nociception. SP-like immunoreactivity is increased peripherally in arthritic joints and centrally in the spinal dorsal horn in adjuvant-induced arthritis (AA) in rats. The release of SP, modulated in vitro by protein kinase C, is also increased. The regulatory mechanisms of SP in the spinal cord are not fully understood in either normal or AA rats. This proposal will test the hypothesis that AA increases the synthesis and/or decreases the degradation of central SP, and modulates SP receptor-mediated events in the spinal cord. The long term goal of this proposal is to elucidate the regulation of the SP-mediated system (SP catabolism, receptor binding and second messenger activity) in the spinal cord and to determine the molecular basis for the increase in SP in the spinal cord during increased afferent impulses associated with chronic pain. The first specific objective is to establish whether the increased content of SP in the spinal cord of rats with AA, and presumed to be in chronic pain, results from its increased synthesis, an enhanced processing of a protein precursor, or a reduction in its rate of degradation. This objective will be accomplished using HPLC/RIA, enzyme assays and molecular biological techniques. The second objective is to characterize functional changes in putative SP receptor(s) in the spinal cord during AA in rats by measuring in vitro SP binding and SP-induced phosphatidylinositol 4,5 bisphosphate (PIP2) hydrolysis. The third objective is to examine the extent to which PKC activity and associated kinetic parameters are modulated in the spinal cord in AA. This will be accomplished by using pharmacological approaches to evaluate the activity of PKC during AA. The fourth objective is to determine the effect of CGRP and/or GAL alone and in combination on SP-mediated PIP2 hydrolysis and on SP metabolism during AA. These studies will help us understand the activity-dependent regulation of SP metabolism, of SP binding, and of the second messengers linked to the SP receptor. This information is essential to the design of efficacious analgesic agents for the management of long term pain such as that associated with arthritis.